Oligonucleotide Nanostructure Markers for Spatial Transcriptomics
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Solution Overview
Problem
Current multi-omics technologies, such as single cell RNA sequencing and spatial profiling methods, face limitations in measuring the whole transcriptome at a single cell level within a tissue section due to insufficient spatial resolution and disruption of cellular interactions during tissue dissociation, which affects molecular layer measurements.
Innovation Solution
A marker system utilizing oligonucleotide nanostructure backbones with attachment sites, labels, and orientation indicators, allowing for reversible attachment and sequencing, enabling efficient tracking and analysis of biological samples in their native context, compatible with both tissue and cell suspension samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If tissue dissociation is performed to enable single cell RNA sequencing and multi-omics measurements, then massively parallel measurements of transcriptome and other data layers can be achieved, but cells are removed from their surrounding taking away signalling inputs and severing attachment to extracellular matrix, which has profound impact on cell morphology and molecular layers
Solution Approach 1:
The invention segments the measurement process into two distinct phases: (1) in situ labeling of cells within intact tissue sections to capture spatial context and cellular interactions, and (2) subsequent dissociation for high-throughput sequencing. This segmentation allows both spatial resolution and massive parallel measurements to be achieved without compromising cellular interaction integrity during the critical labeling phase.
Solution Approach 2:
The invention performs preliminary action by conducting all labeling, marker attachment, and phenotypic characterization on intact tissue sections before dissociation occurs. This ensures that cells are marked while still in their native context with intact signaling inputs and extracellular matrix attachments, thereby preserving the reliability of the measured data even after dissociation for sequencing.
2Reliability
If spatial profiling methods such as Visium or GeoMx are used to maintain tissue context, then cellular interactions are preserved, but neither method can measure the whole transcriptome at the single cell level due to insufficient spatial resolution or plexing capacity
Solution Approach 1:
The invention implements a nested structure where multiple layers of information are encoded within a hierarchical marker system. Primary markers provide spatial location, secondary markers encode transcriptome information, and tertiary markers provide phenotypic data. This nested encoding allows single-cell resolution transcriptome measurement while maintaining spatial context, overcoming the limitations of both Visium (lower resolution) and GeoMx (lower plexing).
Solution Approach 2:
The invention adds another dimension to spatial profiling by implementing multi-layered oligonucleotide encoding that captures not only spatial position but also transcriptome composition and phenotypic states. This multi-dimensional encoding approach enables whole transcriptome measurement at single-cell level within tissue sections, transcending the two-dimensional spatial mapping limitation of conventional methods.
3Adaptability or versatility
If conventional markers are used for marking biological samples, then sample tracking is possible, but a large number of unique markers readable by both microscopy and DNA sequencing cannot be generated
Solution Approach 1:
The invention creates universal markers with multi-functionality that can be detected by multiple modalities. Each marker contains oligonucleotide sequences that enable both fluorescent detection via microscopy and unique identification via DNA sequencing. This universal design allows a single marker system to serve dual purposes: spatial visualization and high-capacity encoding, thereby generating a large number of unique markers without proportionally increasing system complexity.
Solution Approach 2:
The invention exploits parameter changes in oligonucleotide sequences to generate marker diversity. By varying sequence composition, length, and secondary structure of the oligonucleotide markers, a vast number of unique markers can be created that maintain comparable stability and detectability. This parameter-based diversification allows extensive marker libraries to be generated without linearly increasing detection complexity, as the same detection platforms can read diverse sequences.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Enables phenomics-multi-omics characterization at scale by linking phenotypic information from optical readouts to genotyping data, maintaining cellular interactions and morphology, and allowing for the generation of a large number of unique markers readable by microscopy and DNA sequencing.
Implementation Method 1
The attachment oligonucleotide portion of each label includes a unique oligonucleotide sequence configured to bind to a complementary sequence of one of the attachment sites
Data Source
AI summary
A marker for marking a biological sample or a discrete entity including the biological sample includes an oligonucleotide nanostructure backbone with a plurality of attachment sites at predetermined positions, a plurality of labels for attachment to at least one of the attachment sites, and at least a first orientation indicator and a second orientation indicator. Each label includes at least one dye, an encoding oligonucleotide portion configured to encode characteristics of the at least one dye, and an attachment oligonucleotide portion configured to reversibly attach to one of the attachment sites. The attachment oligonucleotide portion of each label includes a unique oligonucleotide sequence configured to bind to a complementary sequence of one of the attachment sites.